Aging causes physiologic changes in gastrointestinal intestinal function that contribute to many age-related disorders. These disorders cause a significant burden both in healthcare costs and quality of life. A shift in the ratio of motor neurons favoring inhibitory (nitrergic) neurons over excitatory (cholinergic) neurons has been observed in the ENS with aging and is thought to play an important role in these disorders. While many have speculated that 'wear and tear' resulting from oxidative stress is the cause for this disturbance, the mechanism(s) underlying the effects of aging on the ENS are poorly understood. Aging is increasingly viewed as a highly regulated process. In the central nervous system (CNS), there is evidence that loss of functional neural stem cells contributes to the aging process and cognitive decline. Similar decline in stem cell function may also be contributing to age-related changes in the ENS. The goal of the proposed research is to study the effect of aging on a population of neural stem cells in the ENS, termed enteric neural stem cells (ENSCs). Recent work by the candidate suggests that ENSCs from old mice differentiate into cells that express higher levels of nNOS, a marker of nitrergic neurons, than those from young mice and that this difference may be due to higher basal Akt activity found in ENS from old mice. The candidate hypothesizes that age-related disturbances in the ENS are due to alterations in ENSC function mediated by the PI3K/Akt/FOXO pathway. The candidate plans to address this hypothesis in two ways. First, the candidate will delineate intrinsic differences in ENSCs isolated by flow cytometry from old and young mice. Specifically, the candidate will perform experiments to evaluate for differences with regards to proliferation, apoptosis, senescence, and differentiation using both in vitro and in vivo (transplantation) approaches. Second, the candidate will investigate the role of the PI3K/Akt/FOXO3 signaling pathway in the aging process in ENSCs. Specifically, the candidate will assess the basal activity of the PI3K/Akt/FOXO3 pathway in old and young ENSCs and evaluate how modulating this pathway through stimulation and inhibition affects their phenotypes. These studies will enhance our understanding of aging in the ENS and lay the foundation for future research including: 1) determining whether caloric-restriction reverses age-related changes to ENSC function, 2) evaluating whether similar age-related changes occur in human ENSCs, and 3) elucidating molecular targets that will reverse the effect of aging on ENSCs. The candidate believes that Stanford, a world-class institution with particular strength in stem cell and aging research, is th ideal environment for developing his academic career. With Dr. Anne Brunet as his senior collaborator, the abundant resources available at Stanford, and a wide range of didactics to broaden his scientific knowledge and technical skills, the candidate is uniquely placed to pursue his goal of becoming an independent investigator.